In vivo, cells integrate a complex set of physical and chemical signals as well as spatial gradients of these signals to maintain proper tissue function. There has been much progress in the artificial replication of these cellular microenvironments by producing substrates with either tailored surface chemistries or regions of varying mechanical compliance. Yet there has been minimal advancement in the design of materials that allow for the independent modulation of both properties to study cellular response. We propose to address this challenge by fabricating artificial extracellular matrix (aECM) films with superimposed gradients and observing cellular behavior on these surfaces. These unique aECM proteins are comprised of cell binding domains of RGD and CS5 and an elastin-derived repeat with a crosslinking site. Gradients in chemical cues can be controlled by the microfluidic mixing of different proteins while gradients in compliance can be achieved by varying the extent of crosslinking. Preparation of this specialty substrate provides a novel experimental approach for examining how competing signals generate a coordinated cellular response. Results from this work can also be applied to create improved materials for biomedical applications.